Chicken erythrocytes are involved in immunity through binding of toll-like receptors (TLRs) with their ligands to activate downstream signaling and lead to cytokine production in erythrocytes. Some avian β-defensins (AvBDs) are constitutively expressed in tissues and some others can be induced by various bacteria and viruses. However, the expression of AvBDs in erythrocytes has not yet been studied extensively. The transcripts of eight AvBDs (AvBD1 to AvBD7, and AvBD9) and liver-expressed antimicrobial peptide-2 (LEAP-2) were found in normal chicken erythrocytes. The expression levels of AvBD2, 4 and 7 were significantly increased (P < 0.01), whereas the levels of AvBD1, 6 and 9 were significantly decreased (P < 0.01) after Marek’s disease virus (MDV) infection. The mRNA expression level of LEAP-2 was not significantly changed after MDV infection. Highest viral nucleic acid (VNA) of MDV in the feather tips among the tested time points was found at 14 days post-infection (d.p.i.). In addition, 35 MD5-related gene segments were detected in the erythrocytes at 14 d.p.i. by transcriptome sequencing. These results suggest that the AvBDs in chicken erythrocytes may participate in MDV-induced host immune responses.

Introduction: It is not clear about mechanisms underlining the inter-segment reassortment of Influenza A viruses (IAVs).We analyzed the viral nucleotide composition (NC) in coding sequences,examined the intersegment NC correlation, and predicted the IAV reassortment using machine learning (ML) approaches based on viral NC features.Methods: Unsupervised ML methods were used to examine the NC difference between human-adapted and zoonotic IAVs. Supervised ML models of random forest classifier (rfc) and multiple-layer preceptor (mlp) were developed to predict the human adaption to IAVs.Results: Our results demonstrated that the frequencies of thymine, cytosine, adenine,and guanine (t, c, a, and g), as well as the content of gc/at were consistently high or low for the segments of PB2, PB1, PA, NP, M1, and NS1 (ribonucleoprotein plus [RNPplus]), between mammalian and avian IAVs or between influenza B viruses (IBVs) and IAVs.RNPplus NC negatively correlated with the NC for HA, NA, and M1 (envelope protein plus [EPplus]). The human-adapted NC accurately discriminated between human IAVs and avian IAVs. A total of 221,184 simulated IAVs with pd09H1N1 EPplus and with RNPplus from other IAV subtypes indicated a high adaption of the RNPplus, from H6N6, H13N2, and H13N8 and other IAVs.Discussion: In summary, there is a distinct human adaption-specific genomic NC between human IAVs and avian IAVs. The intersegment NC correlation constrains segment reassortment. This study presents a novel strategy for predicting IAV reassortment based on viral genetic compatibility.

Transient receptor potential vanilloid 4 (TRPV4) is a calcium-permeable cation channel critical for maintaining intracellular Ca²⁺ homeostasis and is essential in regulating immune responses, metabolic processes, and signal transduction. Recent studies have shown that TRPV4 activation enhances influenza A virus infection, promoting viral replication and transmission. However, there has been limited exploration of antiviral drugs targeting the TRPV4 channel. In this study, we developed the first machine learning model specifically designed to predict TRPV4 inhibitory small molecules, providing a novel approach for rapidly identifying repurposed drugs with potential antiviral effects. Our approach integrated machine learning, virtual screening, data analysis, and experimental validation to efficiently screen and evaluate candidate molecules. For high-throughput virtual screening, we employed computational methods to screen open-source molecular databases targeting the TRPV4 receptor protein. The virtual screening results were ranked based on predicted scores from our optimized model and binding energy, allowing us to prioritize potential inhibitors. Fifteen small-molecule drugs were selected for further in vitro and in vivo antiviral testing against influenza. Notably, glecaprevir and everolimus demonstrated significant inhibitory effects on the influenza virus, markedly improving survival rates in influenza-infected mice (protection rates of 80% and 100%, respectively). We also validated the mechanisms by which these drugs interact with the TRPV4 channel. In summary, our study presents the first predictive model for identifying TRPV4 inhibitors, underscoring TRPV4 inhibition as a promising strategy for antiviral drug development against influenza. This pioneering approach lays the groundwork for future clinical research targeting the TRPV4 channel in antiviral therapies.

(1) Background: With the resurgence of brucellosis epidemics in China in recent years, the chances of a brucella coinfection with other common respiratory pathogens, such as the influenza virus, have increased dramatically. However, little is known about the pathogenicity or the mechanisms of brucella and influenza coinfections. (2) Methods: To clarify the interventions in the early stages of lung damage due to brucella and influenza coinfections, we evaluated the effect of the coinfection on disease progression and mortality using a coinfection model in WT mice and NLRP6^−/− mice, and we verified the function of NLRP6 in infection and proinflammation. (3) Results: The coinfection induced significant respiratory symptoms, weight loss, and a high mortality rate in WT mice. Influenza in the coinfection group significantly increased brucella proliferation in a synergistic manner. Meanwhile, a histological examination showed severe lung tissue destruction and excessive inflammatory responses in coinfected WT animals, and the expression of NLRP6 and IL-18 was dramatically increased in the lung tissues. Furthermore, NLRP6 deletion attenuated lung injuries and inflammation, a reduced bacterial load, and decreased IL-18 protein expression. (4) Conclusions: Our findings indicated that NLRP6 plays a critical role and might be a promising potential therapeutic target for brucella–influenza coinfections.

Influenza viruses cause annual seasonal epidemics and occasional pandemics. Vaccination is most effective at preventing influenza virus infections. Suspensions of the Madin-Darby canine kidney (MDCKs) cell line are used to manufacture cell-based influenza vaccines. In this study, we tested and optimized a range of culture conditions for a chimeric recombinant influenza virus rA/B-H3 in MDCK suspension cells, revealing that the optimal inoculation conditions were multiplicity of infection of 0.001, cell concentration of infection of 4.0 × 10⁶ cells/mL, addition of 2 μg/mL of L-1-tosylamide-2-phenylethyl chloromethyl ketone, time of harvest of 72 h. The optimal parameter setting in a 5-L bioreactor was 50 % dissolved oxygen and pH 7.2 ± 0.05. The rA/B-H3 has stability genetic after 15 passages in MDCKs. The rA/B-H3 had several immune responsesin mice. Hemagglutination inhibition (HAI) antibodies, micro-neutralizing (MN) antibodies, and IgG antibodies were induced in immunized mice, and the mucosal IgA antibody responses were detected in their lung lavage fluids. The IFN-γ-secretion and IL-4-secretion by the mouse splenocytes were induced after stimulation with the specific H3N2 HA protein. Immunized mice resisted the lethal challenge with a wild-type H3N2 influenza virus. This study demonstrated the reliability of the MDCK suspension cell platform for the production of the chimeric H3N2 candidate vaccine, and the rA/B-H3 candidate vaccine is a potentially safe efficial vaccine.

Parainfluenza virus 5 (PIV5), now reclassified as Orthorubulavirus mammalis, is a negative-sense, single-stranded RNA virus belonging to the genus Orthorubulavirus of the family Paramyxoviridae